1. Field of the Invention
The present invention relates to a method of fabrication of a microfluidic device. The present invention also relates to a microfluidic device.
Recent developments in the analytical sciences have focussed on the miniaturisation of separation and detection equipment, mainly for reasons of improved performance and reduced consumption or limited availability of substances. A particular field of interest is that frequently referred to as “lab-on-a-chip”, “microfluidics” or “micro total analysis systems”, which is concerned with the development of instrumentation for the preparation and analysis of chemical or biological samples, the instrumentation having a format that resembles integrated micro-electronic semiconductor circuits. Originally, the developments in this field were aimed at fabrication techniques derived from the micro-electronic field to fabricate miniature separation devices. A major drawback of the systems derived from the fields are needed to establish electro osmotic or electrophoretic principles, which generally can not be sustained on a silicon substrate without electrical breakdown.
2. Brief Description of the Prior Art
Therefore today most of the used microfluidic devices for analysis or synthesis of biological and chemical species are fabricated from two flat electrically insulating glass substrates, with one substrate containing an etched microchannel and drilled or etched access-holes. The glass plates are bonded together so that the microchannel in one substrate forms together with the second glass substrate a microcapillary. In this microcapillary fluids (i.e. liquids and gasses) can be transported or stored, with the intention to perform a chemical reaction between constituents of the fluid, or to separate or mix constituents of portions of the fluid, and subsequently perform chemical or physical analysis on the constituents of the fluid, either on or of the chip. Metal electrodes are frequently integrated on or inserted into these glass chips, such electrodes serving diverse purposes such as electroosmotic or electrokinetic flow control, electrophoretic separation, or electrochemical detection. Ample illustrative examples of such devices can-be found in literature,
D. J. Harrison and co-workers, in: “Capillary electrophoresis and sample injection systems integrated on a planar glass chip”, Analytical Chemistry vol. 64, Sep. 1, 1992, p. 1926, describe a micromachined glass chip, which employs electrokinetic and electroosmotic principles for sample preparation and liquid propulsion, and demonstrate electrophoresis on the chip. An important issue in the fabrication of such glass devices, as well as of devices which comprise one glass substrate and one other substrate, the latter being e.g. a silicon or a polymer substrate, as well as of devices which comprise any combination of these substrate materials, is the sealing of the microfluidic capillary circuit that is formed by combining the two substrates, of which at least one contains an etched or by other means engraved channel pattern.
Some sealing methods use dispensed polymer forming liquids, such as epoxies and the such as, which are considered undesirable for fluidic chip sealing purposes for several reasons, the most important being the difficulties in dispensing a uniformly thick material layer on exact positions along the periphery of an engraved channel, the porosity and mechanical integrity of the material, and the interference of the material with e.g. organic solvents in the channel of the fluidic system during operation.
Other sealing methods are known and summarised below. The methods known for bonding of a glass substrate to a second substrate are inter alia:                1. Deposition of a thin film on one of two glass substrates followed by an anodic (also frequently called electrostatic) bonding process. This metallic or semiconducting layer can be used as intermediate layer. An example of this method is described in the article “Glass-to-glass anodic bonding with standard IC-technology thin films as intermediate layers”, by A. Berthold et. al., Sensors & Actuators A Vol. 82, 2000, pp. 224-228. Described is the use of an intermediate insulator layer such as silicon nitride that acts as a sodium diffusion barrier. An advantage of these anodic bonding methods is that a roughness of several tenths of nanometers can be tolerated without a reduction in bonding quality. Drawback is the high electrical field that is required for the process, which in some cases will result in bonding of channel walls in unwanted locations.        2. Anodic bonding of a glass to a silicon substrate, for example as described in U.S. Pat. No. 3,397,278. Drawback of this method is that it can only be applied for bonding of a glass substrate to a metal or semiconducting substrate, which limits the use of the resulting devices to applications at low electrical fields and relatively low temperatures. The requirement of low temperatures, generally below about 400° C., is the result of the differences in thermal expansion that exist for most combinations of glass and metal or semiconductor substrates, and which lead to unwanted deformations of the substrate sandwich after bonding during temperature cycles.        3. Direct anodic bonding of two insulator substrates, optionally with a metal pattern in-between, as described in U.S. Pat. No. 3,506,424. This method comprises the evaporation of a thin layer of SiO on thin film circuitry, present on a substrate, and subsequent anodic bonding of a glass foil. This procedure results in a sealed electrical connection to the thin film circuitry, which circuitry partially extends to beyond the boundaries of the glass foil. Sealing is achieved because the bonding process presses the glass element on the metal line. This method generally works well for electronic applications, but may lead to unwanted leakage in fluidic applications, in particular if the chip is used at high pressures, which is relevant for separation and synthetic chemistry applications.        4. Thermal glass-to-glass bonding, which consists in heating both substrates to a temperature at which melting starts to occur, or at least to a temperature at which the glass starts to soften, e.g. at 550° C., and pressing the substrates together, by which a bond is formed. This was described in the previously mentioned publication by Harrison et al., and has as important drawbacks the occurrence of leakage when one of the substrates contains surface topography such as metal patterns and the possible deformation of the substrates when they are pressed together in a softened or partially molten state, by which the structural integrity of the fluidic circuit contained in one or both of the substrates will be affected.        5. Bonding of two glass substrates through an intermediate layer of a low-melting-point material, or through an intermediate layer which solidifies from a solution during heat treatment. Such a process is described in the article by H. Y. Wang et al., “Low temperature bonding for microfabrication of chemical analysis systems”, Sensors & Act. B vol. 45, 1997, p. 199-207, in which a spin-on-glass layer is used as an adhesive that solidifies at 90° C. or after one night at room temperature. Drawback of this method is that the layer during dispension or during melting may destroy the structural integrity of the fluidic circuit, due to re-flow of the material.        
Consequently, the previous methods have the disadvantages that an electric field is required for bonding, that a (partially) molten state or application of pressure is required, and/or that the method is limited to a particular choice of substrate material or film material on the substrate.
Further drawbacks of the above methods become evident from the following when sealing is required on metal patterns that are present in-between the two glass plates, between a glass plate and a silicon plate, or between two silicon plates. As discussed by Harrison et al. in the previously mentioned publication, sealing over platinum lines that extended over one of the glass substrates showed liquid leakage even after a careful heat treatment during the thermal bonding procedure. The prevention of leakage is crucial for fluidic microsystems, since leakage will give rise to cross-talk between adjacent fluidic conduits and leads to dead-volumes that give rise to cross-contamination of subsequent sample injections. Leakage is particularly important in fluidic systems which are to be used for gas analysis, systems in which gases are formed by reaction in the channel, or systems in which gas is introduced into a liquid in order to perform a chemical reaction in a chip, such as in the well-known field of microreactors for high-throughput screening of chemical substances.
It is also a requirement to have leak-tight sealing for applications that function with a high pressure inside the fluidic circuit, such as in certain well-known chromatographic methods such as High Performance (High Pressure) Liquid Chromatography (HPLC), HydroDynamic Chromatography (HDC) and some methods of Size Exclusion Chromatography (SEC).
Finally, it is also important to have leak-tight systems whenever the application of the fluidic circuit is in a harsh environment, such as under extremely high pressures or extremely low pressures. High pressures may be present underneath the earth's crust, whereas low pressures or even vacuum may be present in aerospace. Another type of harsh environment is a corrosive environment such as undersea.
One frequently pursued procedure to enhance sealing over metal patterns is that in which a recess is photolithographically defined and etched in one of the substrates, in which subsequently a metal pattern is disposed. Known is a detector integrated with the separation channel, consisting of metal lines that are partially inside and partially outside of the channel, which lines are disposed in the manner using an etched recess in one of the layers. Doing so, a modified electrostatic bonding procedure at a temperature of 350° C. allowed a seal between the layers. This known device is considered undesirable not only because of the extra photolithographic steps that are required during fabrication of the device, but even more because of the necessity of an exact dimensional match and positional alignment of the metal pattern with the etched recess. In particular, the required recess depth uniformity and metal film thickness uniformity over the substrate area, as well as the lithographic overlay quality, is difficult to obtain with most state-of-the-art etching and deposition apparatus, and can only be achieved with very well-tuned and expensive equipment. This is the reason why the method is frequently observed to fail in conventional fabrication environments, and leak-tight sealing is not obtained with the method.